Aggregating protein

[Richard P. Grant]

In article <8pifho$qdb$1 at news.ox.ac.uk>, "James Sandy" 
<james.sandy at pharm.ox.ac.uk> wrote:

> Hi there, I have a problem. I am purifying arylamine n-acetyltransferase 
> and
> the prep is fine until the protein is being concentrated. Around 5 mg/ml 
> the
> protein seems to aggregate and fall out of solution. Does anybody know 
> any
> cunning methods to stop proteins falling out of solution when being
> concentrated?


Hi.  There are, IME, two buggers that cause 'difficult' aggregation (i.e 
changing away from the pI does not help - which of course you should 
try).  The first is proteolytic degradation.  The second is the presence 
of hydrophobic patches on the protein (this how degradation effects 
aggregation).  You can test for the second possibility by keeping the 
prep  at 4C, or adding up to about 33% glycerol or ethylene glycol.  
Both approaches should inhibit hydrophobic interactions.

R

-- 
Richard P. Grant MAD Phil        http://www.gerbil.org.uk/    
Please reply to rpg 'at' mrc-lmb.cam.ac.uk'

         Sex and bugs and rock and roll